The possibility of using polymer nanoparticles for controlled drug/gene delivery at mucosal sites that is extended over several hours to days is expected to lead to effective new therapeutics. However, no such product exists since the human mucosal barrier has been inpenetrable by synthetic nanoparticles, let alone those large enough to carry a sufficient drug payload and slowly deliver it. Trapped particles are typically cleared from mucosal sites over periods of several hours, limiting the ability to achieve controlled drug delivery. Polymer nanoparticles as large as 200-300 nm in diameter have been reported to cross mucus barriers and deliver drugs and genes in rodents. However, although several viruses are known to efficiently cross mucus barriers in humans, no synthetic drug carrier particle has been reported that can move even with modest rates in undiluted human mucus. For example, polymer particles as small as 59 nm, made with polystyrene, have been reported as completely immobile in minimally-diluted human mucus. Using clues from nature (specifically viruses) related to surface chemistry that avoids mucoadhesion, we hypothesize that it is possible to design synthetic nanoparticles that can overcome the human mucus barrier if they possess non- mucoadhesive surfaces. In Aim 1, we will develop nanoparticles of various sizes coated with non- mucoadhesive polymers and we will quantify their efficiency in avoiding entrapment in undiluted human mucus, as well as the average and distribution of individual particle transport rates for hundreds of particles of each size/surface chemistry. In Aim 2, we will treat mucus with agents that alter the pore size of mucus and/or decrease its viscosity and elasticity. We will quantify the effects of these reagents on mucus rheology (e.g., shear-dependent viscous and elastic moduli) using a sensitive strain-controlled rheometer (bulk-fluid rheology) and using nanoparticle tracking rheology (to obtain micro- and macrorheology measurements). We will also quantify the effects of the mucus- altering reagents on the transport rates of particles from Aim 1. Finally, in Aim 3 we will modify a new biodegradable polymer nanoparticle platform to possess appropriate physicochemical properties (based on Aims 1 and 2), and determine if these "real" drug carrier particles are capable of both rapid transport and delivery of potentially useful drugs in human mucus. PUBLIC HEALTH RELEVANCE Mucus is a thick and adhesive material that protects human lungs, GI tract, vagina and other entry points into the body from infection by potentially harmful pathogens. A number of debilitating diseases could be treated more effectively with lower side effects if therapeutics could be slowly delivered at these "mucosal" sites in humans, but current drug delivery systems are incapable of penetrating the mucus barrier. This research seeks to understand the mucus barrier as it relates to prevention of therapeutic particle penetration, and then design new drug carrier particles that can bypass the barrier and provide sustained delivery of a variety of potentially important drugs to treat diseases at mucosal tissues.